Scaffolds for cell collection or elimination

ABSTRACT

A device that includes a scaffold composition and a bioactive composition with the bioactive composition being incorporated therein or thereon, or diffusing from the scaffold composition such that the scaffold composition and/or a bioactive composition captures and eliminates undesirable cells from the body a mammalian subject. The devices mediate active recruitment, sequestration, and removal or elimination of undesirable cells from their host.

RELATED APPLICATIONS

This application is a national stage application, filed under 35 U.S.C.§371, of International Application No. PCT/US2008/007258, filed on Jun.11, 2008, which claims the benefit of provisional application U.S. Ser.No. 60/936,743, filed Jun. 21, 2007, the contents each of which isherein incorporated by reference in its entirety.

SEQUENCE LISTING

The instant application contains a Sequence Listing which has beensubmitted electronically in ASCII format and is hereby incorporated byreference in its entirety. Said ASCII copy, created on Jul. 3, 2017, isnamed 117823-12402 ST25.txt and is 4.0 kilobytes in size.

FIELD OF THE INVENTION

The present invention relates to the field of systemic diseaseprevention, management, and elimination.

BACKGROUND OF THE INVENTION

Some of the most prevalent and debilitating diseases in the US, andthroughout the world, have several fundamental characteristics incommon. First, they can be caused by subpopulations of cells which,through a variety of mechanisms, begin to impair the ability of theirhost to thrive. Second, the degree of severity of disease is closelylinked with the ability of these undesirable cell populations todisseminate throughout multiple physiological systems. The difficultythat medicine faces in treating any condition is selectivity. Theultimate goal of treating disease is to discover a method of eliminatingonly the subpopulations of cells that impair function, whilesimultaneously sparing healthy, uninvolved, cell populations in closeproximity. Many conventional drug and surgical remedies inflict damagingside effects which can either induce a new condition because they areimprecise, and thus inadvertently damage healthy cells, or furtherchallenge the ability of an individual to thrive by making them morevulnerable to future disease.

SUMMARY OF THE INVENTION

The present invention addresses a solution to multiple obstaclesassociated with the in vivo elimination of distinct cell populations.First and foremost, unlike drug remedies, the device is capable ofcapturing, and therefore sequestering, undesirable cells either withinan internal compartment or along its external surface by eitherfiltering cells through pores in the external surface or binding cellsto adhesive proteins along either its internal or external surfaces.This is a critically important innovation, because the mechanismsemployed by the device to eliminate those cells which it contacts areengineered to be highly selective for only targeted cells, allowingsurrounding tissue to remain unaffected. Moreover, this selectivity isparticularly impressive when the device is administered systemically, inwhich case it is capable of first identifying, then capturing, and, ifdesired, eliminating cells that are in the process of spreading diseasewhile preserving epithelial linings and other non-target tissues, aswell as sparing passing blood and immune cells from interaction withtreatments. Furthermore, when the target of the device is a robust celltype that requires a highly toxic agent to induce cell death, theability of this devise to sequester cells away from their hostmicroenvironments is particularly advantageous because concentratingthese cells in a compartment without access to signaling proteins,nutrients, or adequate levels of oxygen itself induces cell death,eliminating the need to introduce a toxic substance into the host.Finally, the device overcomes obstacles such as antibiotic resistanceand drug tolerance by using sequestration, which does not introducecompounds systemically, as well as a bioactive composition that includesmolecules or cells capable of activating signaling pathways within thetarget cell population that will lead to self-destruction.

This device includes a scaffold composition which incorporates or iscoated with a bioactive composition; the device attracts, adheres,captures and eliminates targeted undesirable cells. The device executesthese functions by a variety of methods that include or exclude the useof a bioactive composition. Depending on the application for which thedevice is used, the device regulates capture and survival through thephysical or chemical characteristics of the scaffold itself. Forexample, the scaffold composition is differentially permeable, allowingcell passage only in certain physical areas of the scaffold. Thepermeability of the scaffold composition is regulated, for example, byselecting or engineering a material for greater or smaller pore size,density, polymer cross-linking, stiffness, toughness, ductility, orviscoelasticity. The scaffold composition contains physical channels orpaths through which cells contact a specific bioactive compositionwithin the device or move into an intended compartment within thedevice. The scaffold composition is optionally organized intocompartments or layers, each with a different permeability, so thatcells are sorted or filtered in order to exclude targeted subpopulationsfrom accessing or participating in the host environment whilesimultaneously allowing non-targeted cell types to pass through thedevice unaltered. Migration of target cell populations through thedevice is also regulated by the degradation, de- or re-hydration,oxygenation, chemical or pH alteration, or ongoing self-assembly of thescaffold composition. These processes are driven by diffusion orcell-secretion of enzymes or other reactive chemicals. Following theircapture, resident cells are prohibited from accessing the hostenvironment, which provides necessary means for survival including hostcell contact, signaling molecules, sustaining nutrients, and adequatelevels of oxygen. Thus, having been removed from the context of anybiological niche, captured cells are sequestered or eliminated. Once thecaptured population is removed the scaffold composition either degradesor is collected from the subject.

Exemplary cell types that are targeted for sequestration or quarantineby this device, and are eliminated by environmental deprivation alone,and without the aid of a bioactive composition, include circulatingmicroorganisms such as bacteria, viruses, parasites, worms, protozoa, aswell as mobile mammalian tissues including immune cells, metastaticcancer cells, sickle-shaped or damaged blood cells, and epithelial cellsthat have lost intercellular contacts.

Optionally, a bioactive composition with means to physically bind andadhere target cells to the scaffold composition is added to ensure thatcells do not escape from the scaffold once they have contacted it.Exemplary components of such an adhesive bioactive composition areeither found in nature or engineered, and are listed below alongside thecell types for which they are specific.

Alternatively or in addition, the processes of cell attraction,adhesion, capture, or elimination are regulated primarily by one or morebioactive compositions. By varying the concentration of attractivemolecules, the bioactive composition draws mobile or circulating cellsinto the device. The following are examples of attractive compounds thatare used alone or in combinations to attract mobile cells to thescaffold composition: multivalent acrylamide polymers or polylysinetagged with either a sugar (galactose, fucose, ribose, or anyderivatives thereof) or an amino acid (aspartate, leucine, serine) toattract bacteria; cytokines or chemokines to attract immune andmetastatic cancer cells. By including particular subsets of adhesionmolecules, the bioactive composition serves a selectivity function,second to the sorting and filtering function performed by the scaffoldcomposition itself, wherein adhesion molecules that preferentially bindtarget cell types are incorporated into the device to differentiatebetween morphologically or physically similar target and non-target cellpopulations that might otherwise pass through porous filters. Forexample, the adhesion molecules including, but not limited to, thefollowing are incorporated into bioactive compositions with means tocapture microorganisms: adhesins to bind prokaryotes; polysaccharides tobind bacteria; sugar sialic acid, capsid-binding proteins, viralattachment proteins, isolated plasma membrane fractions from cellulartargets to bind viruses. Alternatively adhesion molecules including, butnot limited to, the following are incorporated into bioactivecompositions to capture eukaryotic, or mammalian, cells: integrins, Igsuperfamily members, cadherins, or selectins.

Unique combinations of attraction or adhesion molecules incorporatedinto the scaffold composition to differentially capture target cellpopulations, wherein target cell capture is either permanent ortemporary depending upon which duration is most advantageous for theintended use of the device. For example, adhesive interactions withmicroorganisms can either be permanent or temporary depending upon thefunction of the device. If the device is used as a diagnostic tool forwhich it will be collected, semi-permanent, strong but temporary,adhesive interactions are preferred because the microorganisms can notescape from the device in the host, but can be separated from the deviceupon collection to allow for the isolation and identification of thecaptured cells by a laboratory technician. However, if the device isused as a treatment for a known infection, a permanent adhesiveinteraction is preferred because the microorganisms are sequestered andkilled while attached to the scaffold composition and do not need to berecovered. Alternatively, the scaffold itself can be dissolved in vitrofollowing collection to release microorganisms or mammalian cellsadhered to the bioactive composition for further analysis.

By varying the components of the bioactive composition that lead to celldeath, the device eliminates cells efficiently with minimal disturbanceto non-targeted cell types. The device employs two primary mechanisms.By the first mechanism, target cell populations are attracted to orselectively filtered through the scaffold composition where they bindadhesive agents incorporated into the scaffold. These captured cells arethen eliminated by a variety of mechanisms, all of which act exclusivelywithin the device. By the second mechanism, the device is placed in aspecific location where a target cell population resides, where itcontacts neighboring cells and is anchored, e.g., adheres, in order toprevent diffusion or migration of the device. In this scenario, theclosely juxtaposed cells are exposed to particles, compounds, orengineered cells that are released or allowed to diffuse from thedevice, disseminating only a short distance. Target cell specificity, inthis second case, is achieved by the differential placement of thedevice, the specific compounds released, or manipulation of cell typeshoused within the device allowing them only to attack cells having acharacteristic morphology or surface protein expression profile.

The following descriptions elucidate specific means by which target cellpopulations are eliminated either following capture and sequestrationinto the scaffold composition, or following release of particles,compounds, drug agents, or engineered cell types into the tissuesurrounding an implanted device.

Antibiotic compounds including, but not limited to, amikacin,gentamicin, kanamycin, neomycin, netilmicin, streptomycin, tobramycin,teicoplanin, vancomycin, azithromycin, clarithromycin, clarithromycin,dirithromycin, erythromycin, roxithromycin, troleandomycin, amoxicillin,ampicillin, azlocillin, carbenicillin, clozacillin, dicloxacillin,flucozacillin, mezlocillin, nafcillin, penicillin, piperacillin,ticarcillin, bacitracin, colistin, polymyxin B, ciprofloxacin, enoxacin,gatifloxacin, levofloxacin, lomefloxacin, moxifloxacin, norfloxacin,oflazacin, trovafloxacin, mafenide, sulfacetamide, sulfamethizole,sulfasalazine, sulfisoxazole, trimethoprim, cotrimoxazole,demeclocycline, soxycycline, minocycline, oxytetracycline, ortetracycline are incorporated, alone or in any combination thereof, intoa bioactive composition within devices used for treatment of bacterialinfection.

Antiviral compounds including, but not limited to, amantadine,rimantadine, pleconaril, acyclovir, zidovudine, lamivudine, fomivirsen,zanamivir, oseltamivir, or interferon alpha are incorporated, alone orin any combination thereof, into a bioactive composition within devicesused for treatment of viral infection.

Antiparasitic compounds including, but not limited to, ivermectin,diethylcarbamazine, eflornithine, melarsoprol, pentamidine, suramin,pentavalent antimonials, liposomal amphotericin B, miltefosine,chloroquine, sulphadoxine, mefloquine, artemisinins, artemether,lumefatrine, atovaquone, proguanil, chlorproguanil, dapsone,fosmidomycin, DB289, pyrimethamine, cycloguanil, sulphamethoxazole,difluoromethylornithine, antifungal trizoles (for example,posaconazole), risedronate, levamisole, or albendazole, areincorporated, alone or in any combination thereof, into a bioactivecomposition within devices used for treatment of parasitic, protozoan,or worm-derived infections.

Antifungal compounds including, but not limited to, sertaconazole,sulconazole, tioconazole, fluconazole, itraconazole, isavuconazole,ravuconazole, posaconizole, voriconazole, terconizole, terbinafine,amorolfine, naftifine, butenafine, anidulafungin, caspofungin,micofungin, ciclopirox, flucytosine, griseofulvin, haloprogin,tolnaftate, undecylenic acid, or ampho B lipid, are incorporated, aloneor in any combination thereof, into a bioactive composition withindevices used for treatment of fungal infections.

Mammalian cell types are targeted and eliminated by the device. Toeliminate captured cells the device utilizes a variety of mechanisms.Bioactive compositions incorporated into the scaffold composition withmeans to destroy captured cells include, but are not limited to,phagocytic cells with or without compounds having means to augment theiractivity; engineered viruses; humanized monoclonal antibodies toactivate the immune system, bind and block receptor activity, orspecifically deliver a toxin with which it is bound; pro-apoptoticligands, such as FasL, TNF, TRAIL, and caspase-activators; radioactiveisotopes, such as Bismuth-213 Holmium-66, Iodine-125, Iodine-131, andLutetium-177; toxins, including hemotoxins, necrotoxins, andneurotoxins; and chemotherapy agents.

To accomplish long-lasting weight-loss for individuals who are unable tocontrol their fat intake or storage by any other method, and to avoidrisks associated with surgical interventions, one or more device(s) areimplanted on or around adipose tissue, usually located either beneaththe skin (subcutaneous) or around internal organs (intraperitonealcavity), wherein one or more bioactive compositions conjugated to thescaffold composition adhere adipocytes or pre-adipocytes (precursorsthat give rise to adipocytes) and induce cell death following contactwith either the scaffold composition or a diffusible bioactivecomposition released from the device. The abundance and placement ofimplanted devices controls the degree to which adipocytes andpre-adipocytes are targeted, although ultimately, some tissue remains,as these cells are only undesirable when overly expanded or abundant,particularly when they reside in close proximity to and inhibit thefunction of vital internal organs. White adipose tissue (WAT) istargeted by the device because the alternate type, brown adipose tissue,performs functions essential for the health of the subject. Exemplarysurface proteins that are used by the bioactive composition tospecifically recognize immature and mature white adipocytes include, butare not limited to, serum amyloid A (SAA), adipophilin, and/orlipoprotein lipase (LPL). Adipocytes or their precursors, are eliminatedby induction of apoptosis via a bioactive composition released from astationary, and therefore implanted, device.

Neoplastic cell types, malignant or benign, are targeted and eliminatedby the device. Cells that contribute to the formation of malignantneoplasms are targeted as a treatment for cancer while those cells thatcontribute to benign neoplasms are targeted as a preventative measure todecrease the future risk that those neoplasms, often referred to as “insitu tumors,” could acquire traits consistent with a malignantformation. One characteristic shared between all neoplasms, regardlessof their potential to cause cancer, is a population of rapidly dividingcells. Therefore, to remove neoplasms from the host, the device isimplanted in the area of the tumor, either in or around the cell mass,where one bioactive composition containing adhesive compounds, listedabove for mammalian cells, mediates binding of tumor tissue to thescaffold, and any number of additional bioactive compositions comprisingcompounds or drugs with means to prevent or arrest cell division contactthe captured cells in situ or diffuse from the scaffold composition intothe surrounding tissue. Exemplary compounds with means to slow or retardthe division of cells within a neoplasm include, but are not limited to,Dacarbazine/DTIC, Fluorouracil/5-FU, Fludarabine, Gemcitabine,Trastuzumab/Herceptin, Hydroxyurea/Hydrea, Idarubicin, Ifosfamide,Irinotecan, Cladribine/Leustatin, Mercaptopurine/Purinethol/6-MP,Methotrexate, Mithramycin/Plicamycin, Mitomycin,Mitoxanthrone/Novatrone, Navelbine/Vinorelbine, Nitrogen Mustard,Rituxan, Paclitaxel/Taxol, Docetaxel/Taxotere, Topotecan,Velban/Vinblastine, Vincristine, or Etoposide/VP-16 are incorporated,alone or in any combination thereof, into a bioactive composition withindevices used for reversal of tumor formation. Cancers for which thedevice is used as a treatment or preventative measure at any point inthe life span of the subject include, but are not limited to, acutelymphoblastic leukemia, acute myeloid leukemia, adrenocorticalcarcinoma, AIDS-related cancer, AIDS-related lymphoma, anal, appendix,cerebellar astrocytoma, cerebral astrocytoma, basal cell carcinoma, bileduct, bladder, one, osteosarcoma, malignant fibrous histiocytoma, brainstem glioma, brain, malignant glioma, ependymoma brain, medulloblastoma,supratentorial primitive neuroectodermal tumor, visual pathway andhypothalamic glioma, breast, bronchial adenomas, bronchial carcinoids,Burkitt's lymphoma, carcinoid tumor, gastrointestinal carcinoid tumor,carcinoma of unknown primary, central nervous system lymphoma, cervical,chronic lymphoid leukemia, chronic myelogenous leukemia, chronicmyeloproliferative disorders, colon, colorectal, cutaneous T-celllymphoma, mycosis fungoides, Sezary syndrome, endometrial, ependymoma,esophageal, Ewing's family of tumors, extracranial germ cell tumor,extragonadal germ cell tumor, extrahepatic bile duct, interocularmelanoma, retinoblastoma, eye, gallbladder, gastric (stomach),gastrointestinal carcinoid tumor, gastrointestinal stromal tumor,ovarian germ cell tumor, germ cell tumor, gestational trophoblastictumor, glioma, hairy cell leukemia, head, neck, hepatocellular,Hodgkin's lymphoma, hypopharangeal, islet cell carcinoma (endocrinepancreas), Kaposi's sarcoma, kidney (renal cell), kidney, laryngeal, lipand oral cavity, liver, lung (small cell), lung (non-small cell),Non-Hodgkin's lymphoma, Waldenstrom's macroglobulinemia, melanoma,merkel cell carcinoma, mesothelioma, metastatic squamous neck cancerwith occult primary, mouth, multiple endocrine neoplasia syndrome,multiple myeloma, plasma cell neoplasm, myelodysplastic syndromes,myeloproliferative diseases, nasal cavity, paranasal sinus,nasopharyngeal, neuroblastoma, oral, oral cavity, oropharyngeal,ovarian, ovarian epithelial, ovarian low malignant potential tumor,pancreatic, parathyroid, penile, pharyngeal, pheochromocytoma,pineoblastoma, pituitary, pleuropulminary blastoma, prostate, rectal,renal pelvis and uterer, transitional cell cancer, rhabdomyosarcoma,salivary gland, soft tissue sarcoma, uterine sarcoma, skin(non-melanoma), small intestine, squamous cell carcinoma, T-celllymphoma, testicular, throat, thymoma, thymoma carcinoma, thymiccarcinoma, thyroid, unknown primary site cancer, unknown primary sitecarcinoma, urethral, uterine, vaginal, vulvar, or Wilm's tumor.

To prevent metastatic cells from escaping their primary tumor sites anddisseminating to remote anatomical locations, one or more device(s) isimplanted in or around malignant neoplasms. A bioactive composition isincorporated into or onto the scaffold composition, e.g. the scaffoldnot only contains cell-death-inducing agents, but also anti-metastaticcompounds. Tumor cells that are preparing to enter the blood streamexpress cytokine receptors through which cytokines or chemokines providemigrational encouragement and instructions. Thus, bioactive compositionswith means to retard or arrest metastasis include, but are not limitedto, the following inhibitors of chemokine receptor activity: AMD3100,T22, TN14003, T140, TC14012, 4F-bTE, BX-471, Met-CCL5, TAK779, TAK220.By inhibiting chemokine receptor function, the preceding compoundsprevent tumor cells from reacting to pro-metastatic or migratory cuespresent in their environment.

To capture mobile or circulating cancer cells which have broken free oftheir primary tumor formations, the device is administered systemicallywherein it circulates in the blood, gastrointestinal tract, lymphaticfluid, or cerebral spinal fluid, and adheres and traps cancer cells byadhering to surface proteins unique to metastatic cell lines. Uponcapturing these cells, the device is either used as a diagnostic tool,collected to identify the captured cells and determine the site of theprimary tumor and metastasis. Alternatively, the device is used as atreatment method, and therefore, not collected, but rather a bioactivecomposition is incorporated within the device to eliminate the capturedcell population. When the device is used to trap and kill metastaticcancer cells, and does not require collection, the scaffold compositiondegrades or is physically removed from the body, e.g. surgicallycollected. Alternatively, the device is used as an arterial stent. Inthis embodiment, the device is implanted and acts as a sieve, allowingall cell types to pass through, except for metastatic cancer cells thatare captured by incorporating into the scaffold composition a bioactivecomposition with means to specifically adhere these cells. Uponcontacting the stent device, captured cells are either instructed byeither the flow dynamics inside of the scaffold or by a second bioactivecomposition to either enter an inside chamber where these cells die, orthe captured cells are killed by direct interaction with a component ofa second bioactive composition incorporated into the framework of thestent. Finally, a combination of these mechanisms is used to trap andkill metastatic cancer cells that either contact the framework or innerchambers of the scaffold composition. Furthermore, the bioactivecompositions incorporated into either a mobile device or stent-likedevice can be altered to specifically adhere cells that contribute toforms of leukemia and lymphoma.

Autoimmune disease can be prevented, managed, or treated using thedevice. For this purpose the device is administered either locally orsystemically and incorporated with a bioactive composition with means toirreversibly adhere immune cells predisposed to recognize self-antigensas “foreign,” and in response to these antigens, signal to or recruitother immune cells, proliferate, or attack host tissues. A bioactivecomposition is incorporated into the scaffold composition that includesself-antigens, isolated from tissue samples taken from the subject, totarget autoimmune cells, as well as, adhesive proteins to bind thoseimmune cells, and signaling proteins to either induce cell death orprevent further immune responses. Ultimately, the device capturesautoimmune cells by providing an alternative target for these immunecells as a way to competitively inhibit host antigen binding.

The device targets, captures, and eliminates malfunctioning host cellsthat cause damage to the subject by means of failing to properly performa given function or by accumulating within the host, interrupting thefunction of healthy cells. For example, the device is used topreemptively capture and remove clot-forming red blood cells as a directresult of Sickle Cell Disease. Individuals with this condition expressan abnormal form of hemoglobin, called hemoglobin S (named forhemoglobin “sickle”), on the surface of red blood cells. The presence ofhemoglobin S induces several morphological changes: red blood cellsdevolve from soft, round cells that can fit through small-diametervessels into hardened, sickle-shaped cells, which accumulate at pointsalong the circulatory path, ultimately decreasing the flow of blood tonearby tissues. Moreover, blood cells that express hemoglobin S have asignificantly shorter life-span than cells that express the wild typeform. The device is administered intravenously to a subject as either amobile device, or an arterial stent, both of which filter blood. Sicklecells are identified and trapped either by virtue of their alteredmorphology or by binding to hemoglobin S, thereby adhering these cellsto the scaffold composition. Following their capture, sickle cells areinduced to die prematurely via pro-apoptotic ligands or they areengulfed by phagocytic cells belonging to a bioactive composition. Bothmethods of sickle cell elimination occur within the confines of thescaffold composition.

The device is used to aid in the treatment of peripheral and centralnerve injury. When a nerve is transected, such that the axons of nervecells comprising the nerve bundle are severed, supporting cells, calledglial cells, respond by forming a “glial scar” at the site of injury.The formation of this scar prevents the axon from regenerating bycreating a physically impenetrable barrier between the cell's body andits synaptic targets. To prevent the glial scar from forming, the deviceis placed at the site of nerve transaction and throughout thesurrounding tissue wherein bioactive compositions incorporatedthroughout the device adhere glial cells to the scaffold composition,capturing them, and/or eliminating them, in order to prevent theirmigration to or proliferation at the site of injury. When used to treatinjuries sustained to the central nervous system, in which nerveregeneration is also chemically prohibited, bioactive compositionsincorporated into the scaffold composition further comprise compoundswith means to chelate anti-regeneration ligands (such as, Nogo), blockanti-regeneration receptors (the Nogo receptor, Nogo-R), and thosecompounds, such as growth factors, with means to promote axonelongation. Bioactive compositions with means to stimulate nerve growthand/or attenuate anti-regeneration cues are either bonded to thescaffold composition or allowed to diffuse into the surrounding tissueor space.

To eliminate organic accumulations, plaques, or waste products thatimpair host function, the device contains live cell types or compoundsthat degrade these undesirable compositions. Exemplary cell types thatclear harmful debris or accumulations commonly belong to the immunesystem and include, but are not limited to, macrophages, neutrophils, ornatural killer (NK) cells. These immune cells are engineered tospecifically target cell populations when they are released from thescaffold composition, however, these cells also reside inside the deviceand clear debris from the scaffold composition as captured cell typesare eliminated. The presence of these cells allows the device tocontinually trap cells. To activate resident phagocytic cells,adiponectin, a protein normally produced by adipocytes and known tofacilitate phagocytosis by forming a bridge between dead cells andmacrophages, is further incorporated into one or more of the bioactivecomposition(s).

Genetically-manipulated viruses that are engineered to specificallyattack target cell populations are incorporated into one or morebioactive composition(s) of the devise. These engineered viruses eitherpermanently reside within the confines of the scaffold composition or,alternatively, diffuse into surrounding tissues following implantationof the device, dependent upon the precision of their specificity and thephysical characteristics of the device placement. For example, oncolyticviruses are engineered to specifically infect cancer cells either bymodification of their coat protein specificity or by altering the genomeof the virus to render it replication-incompetent in any cell type otherthan a cancer cell. Adenovirus and herpes simplex virus are often usedto infect cancer cells and are included in the bioactive composition ofthe device. Alternatively, some oncolytic viral strains are specific forcancer cells either due to the increased susceptibility of most cancercells, as in the case of rhabdovirus, or, as a result of their naturalspecificity, as in the case of poliovirus, which evolved to be aneuropathogen. Furthermore, these viruses kill tumor cells either bylysing the cell, which is a normal function of a virus, or by deliveringpro-apoptotic and/or anti-angiogenic genes to induce cell death, whichis a genetically engineered function of these viruses. To prevent thehost immune system from recognizing engineered viruses and inducing aresponse that could eliminate them before they have executed theirfunction, e.g. infect cancer cells, these viruses are coated with apolymer, such as polyethylene glycol, which serves as a shield toantibodies.

The device controls and directs the migration of cells through itsstructure. Chemical affinities are used to channel cells towards aspecific area of contact or quarantine. For example, attraction andadhesion molecules are used to attract or retard the migration of cells,respectively. By varying the density and mixture of those bioactivesubstances, the device controls the specificity of capture and locationof adhesion. The density and mixture of these bioactive substances iscontrolled by initial doping levels or concentration gradient of thesubstance, by embedding the bioactive substances in scaffold materialwith a known leaching rate, by release as the scaffold materialdegrades, by diffusion from an area of concentration, by interaction ofprecursor chemicals diffusing into an area, or by production/excretionof compositions by resident support cells. The physical or chemicalstructure of the scaffold also regulates the diffusion of bioactiveagents through the device.

The device is composed of a biocompatible material. One of the mostcommon embodiments of the device incorporates the extracellular matrix(ECM) as a component of one or more bioactive composition(s) in order toadhere a wide variety of foreign and host cell types. In this case, theECM is chemically cross-linked to the scaffold composition. Regardlessof the tissue of origin, ECM components generally include three generalclasses of macromolecules: collagens, proteoglycans/glycosaminoglycans(PG/GAG), and glycoproteins, e.g., fibronectin (FN), laminin, andthrombospondin. ECM components associate with molecules on the cellsurface and mediate adhesion and/or motility. Preferably, the ECMcomponent associated with the scaffold is a proteoglycan attachmentpeptide or cyclic peptide containing the amino acid sequencearginine-glycine-aspartic acid (RGD). Proteoglycan attachment peptidesare selected from the group consisting of G₄RGDSP (SEQ ID NO: 1), XBBXBX(SEQ ID NO: 2), PRRARV (SEQ ID NO: 3), YEKPGSPPREVVPRPRPGV (SEQ ID NO:4), RPSLAKKQRFRHRNRKGYRSQRGHSRGR (SEQ ID NO: 5), and RIQNLLKITNLRIKFVK(SEQ ID NO: 6), and cell attachment peptides are selected from the groupconsisting of RGD, RGDS (SEQ ID NO: 12), LDV, REDV (SEQ ID NO: 13), RGDV(SEQ ID NO: 14), LRGDN (SEQ ID NO: 7), IKVAV (SEQ ID NO: 8), YIGSR (SEQID NO: 9), PDSGR (SEQ ID NO: 10), RNIAEIIKDA (SEQ ID NO: 11), RGDT (SEQID NO: 15), DGEA (SEQ ID NO: 16), and VTXG (SEQ ID NO: 17).

Components of the ECM, e.g., FN, laminin, and collagen, interact withthe cell surface via the integrin family of receptors, a group ofdivalent cation-dependent cell surface glycoproteins that mediatecellular recognition and adhesion to components of the ECM and to othercells. Ligands recognized by integrins typically contain an RGD aminoacid sequence that is expressed in many ECM proteins. Exemplarymolecules that mediate cell adhesion and/or movement include FN,laminin, collagen, thrombospondin 1, vitronectin, elastin, tenascin,aggrecan, agrin, bone sialoprotein, cartilage matrix protein,fibronogen, fibrin, fibulin, mucins, entactin, osteopontin, plasminogen,restrictin, serglycin, SPARC/osteonectin, versican, von WillebrandFactor, polysaccharide heparin sulfate, cell adhesion moleculesincluding connexins, selectins, collagen, RGD (Arg-Gly-Asp) and YIGSR(Tyr-Ile-Gly-Ser-Arg) (SEQ ID NO: 9) peptides, glycosaminoglycans(GAGs), hyaluronic acid (HA), integrins, selectins, cadherins andmembers of the immunoglobulin superfamily. Carbohydrate ligands of theECM include the polysaccharides hyaluronic acid, andchondroitin-6-sulfate.

To create a multifunctional device, a plurality of bioactivecompositions are covalently linked to the scaffold composition, keepingthese compositions relatively immobilized in or on the scaffoldcomposition. In other cases, additional bioactive compositions arenoncovalently associated with the scaffold. Noncovalent bonds aregenerally one to three orders of magnitude weaker than covalent bondspermitting diffusion of the factor within the compartments of thedevice, and ultimately, out of the scaffold and into surroundingtissues. Noncovalent bonds include electrostatic, hydrogen, van derWaals, π aromatic, and hydrophobic interactions.

The scaffold composition is also contacted with a second bioactivecomposition. The second bioactive composition is sometimesnon-covalently associated with the scaffold composition to yield alayered scaffold, i.e., a scaffold composition that includes one or morebioactive substances. The application of contacting of the scaffoldcomposition with a second bioactive composition is optionally repeatedto yield a plurality of scaffold structures, e.g., each of thecontacting steps is characterized by a different amount of the secondbioactive composition to yield a gradient of the second bioactivecomposition in the scaffold device. Rather than altering the amount ofcomposition, subsequent contacting steps involve a different bioactivecomposition, i.e., a third, fourth, fifth, sixth . . . , composition ormixture of compositions, that is distinguished from the priorcompositions or mixtures of prior doping steps by the structure orchemical formula of the factor(s). The layers, or components of thescaffold form a gradient to lure undesirable cells into the device toone another and/or insertion of semi-permeable, permeable, orimpermeable membranes within or at one or more boundaries of the deviceto further controls/regulate the locomotion of cells or bioactivecompositions into the scaffold to trap the cells therein, as well as theelimination of resident cell populations.

The scaffold composition can be constructed using a variety of materialsthat will be chosen according to its desired function. In all cases, thescaffold composition is biocompatible, meaning that the scaffoldcomposition itself does not induce damage or activate the host immunesystem. The composition is bio-degradable/erodable or resistant tobreakdown in the body. Relatively permanent (degradation resistant)scaffold compositions include metals and some polymers such as silk.

The scaffold composition is used for a variety of functions for whichone or more of the bioactive compositions destroy(s) the captured cellpopulation. Therefore, the scaffold itself does not need to existindefinitely either internal or external to the host, and is composed ofa biodegradable material. Breakdown of the scaffold composition as aresult of any or all of the following events: the captured cells releaseone or more corrosive compound(s), the local host environment where thescaffold resides contains one or more corrosive compound(s), the processof eliminating the captured cells produces one or more corrosivecompound(s), the bioactive composition itself compromises the structuralintegrity of the scaffold, the scaffold disassembles upon collection. Inone example, cells mediate degradation of the scaffold matrix, i.e., thescaffold composition is enzymatically digested by a composition elicitedby a resident cell. In this case, polymer main chains or cross-linkscontain compositions, e.g., oligopeptides, that are substrates forcollagenase or plasmin, or other enzymes produced by within or adjacentto the scaffold.

For some applications, the scaffold composition preferably degrades at apredetermined rate based on a physical parameter selected from the groupconsisting of temperature, pH, hydration status, and porosity, thecross-link density, type, and chemistry or the susceptibility of mainchain linkages to degradation or it degrades at a predetermined ratebased on a ratio of chemical polymers. For example, a high molecularweight polymer comprised of solely lactide degrades over a period ofyears, e.g., 1-2 years, while a low molecular weight polymer comprisedof a 50:50 mixture of lactide and glycolide degrades in a matter ofweeks, e.g., 1, 2, 3, 4, 6, 10 weeks. A calcium cross-linked gelscomposed of high molecular weight, high guluronic acid alginate degradeover several months (1, 2, 4, 6, 8, 10, 12 months) to years (1, 2, 5years) in vivo, while a gel comprised of low molecular weight alginate,and/or alginate that has been partially oxidized, will degrade in amatter of weeks.

Exemplary scaffold compositions include polylactic acid, polyglycolicacid, PLGA polymers, alginates and alginate derivatives, gelatin,collagen, fibrin, hyaluronic acid, laminin rich gels, agarose, naturaland synthetic polysaccharides, polyamino acids, polypeptides,polyesters, polyanhydrides, polyphosphazines, poly(vinyl alcohols),poly(alkylene oxides), poly(allylamines)(PAM), poly(acrylates), modifiedstyrene polymers, pluronic polyols, polyoxamers, poly(uronic acids),poly(vinylpyrrolidone) and copolymers or graft copolymers of any of theabove. One preferred scaffold composition includes an RGD-modifiedalginate.

For some applications, porosity of the scaffold composition influencesselectivity of the cells captured by or permitted through the device.Pores are nanoporous, microporous, or macroporous. For example, thediameter of nanopores are less than about 10 nm; micropore are in therange of about 100 nm-20 μm in diameter; and, macropores are greaterthan about 20 μm (preferably greater than about 100 μm and even morepreferably greater than about 400 μm). In one example, the scaffold ismacroporous with aligned pores of about 400-500 μm in diameter.

The devices are manufactured in their entirety in vitro. Furthermore thedevices can be assembled around or in contact with engineered culturedcells or viruses with means to attract, adhere, or eliminate capturedtarget cells, in order to produce a scaffold and bioactive compositioncombination that includes living or dead cells. Alternatively, thedevice is manufactured in two or more (3, 4, 5, 6, . . . 10 or more)stages in which one layer or compartment is made and coated withbioactive composition followed by the construction of a second, third,fourth or more layers, which are in turn coated with bioactive materialin sequence. Each layer or compartment is identical to the others ordistinguished from one another by the number, genotype, or phenotype ofthe seed cell population as well as distinct chemical, physical andbiological properties of the scaffold or bioactive composition.

Therapeutic applications of the device include diagnosis of infection bymicroorganisms, or the discovery and diagnosis of diseased host tissuesthat contribute to sickle cell disease, leukemia, metastatic cancer,autoimmune conditions, or inflammatory disease. Moreover, this device isused as a diagnostic tool to capture and identify cancer cells that havemetastasized potentially from multiple primary tumors, therebyeliminating the need for multiple biopsies or more invasive surgicaloptions. Alternatively, this devise is used as a medical treatment toeliminate microorganisms, malignant neoplasms, leukemia, lymphoma,arterial blockages, adipose tissue, inflammation, glial scars, andprotein deposits. Furthermore, this devise can be used for preventativemedicine to assess exposure to microorganisms, eliminate benignneoplasms, eliminate immune cells that activate to self-antigens, aswell as prevent arterial build-up or glial scaring following nervedamage.

Other features and advantages of the invention will be apparent from thefollowing description of the preferred embodiments thereof, and from theclaims. References cited are hereby incorporated by reference.

DETAILED DESCRIPTION OF THE INVENTION

The device contains a scaffold composition incorporated with one or morebioactive compositions. The most fundamental functions of the device arecapturing and eliminating target cells from a host organism. In thiscase, the “host” is any mammalian subject and the term “host cell”describes any cell that is recognized by a healthy immune system asdisplaying self-antigens. Alternatively, a “foreign cell” is meant todescribe any cell that is recognized by a healthy immune system asdisplaying non-self, or foreign, surface antigens as well as any cellthat was not born, or generated, within the host organism. The term“undesirable” cell is meant to describe any cell that increases the riskor susceptibility of the host to infection or disease, or any cell thatdecreases the ability of the host to thrive. The term “scaffoldcomposition” is meant to describe a biocompatible, biodegradable ornon-biodegradable frame, used to deliver a bioactive composition to aparticular location of the body, or to create a physical niche into oronto which targeted cell types accumulate and, by this act, aredestroyed or removed from the context of the host body. The term,“bioactive composition” is meant to describe any combination ofmolecules, compounds, proteins, or cell types having means to alter thebehavior of the target cell type. The bioactive compositions used withinthe invented device either remain incorporated with the scaffoldcomposition, or they may diffuse from the scaffold structure to contactsurrounding target tissues.

Resident cells, incorporated into a bioactive composition prior toadministering the device to a subject, including immune cells andviruses, are optionally genetically manipulated by the introduction ofexogenous genetic sequences or the inactivation or modification ofendogenous sequences. For example, recombinant genes are introduced tocause the cells to encode or make proteins that are otherwise lacking inthe host or target tissue. Production of scarce but desirable proteins(in the context of certain tissues) is augmented by transplantinggenetically engineered cells. General mammalian cell culture techniques,cell lines, and cell culture systems are described in Doyle, A.,Griffiths, J. B., Newell, D. G., (eds.) Cell and Tissue Culture:Laboratory Procedures, Wiley, 1998, the contents of which areincorporated herein by reference.

Scaffold Compositions and Architecture

Components of the scaffolds are organized in a variety of geometricshapes (e.g., beads, pellets), niches, planar layers (e.g., thinsheets). For example, multi-component scaffolds are constructed inconcentric layers each of which is characterized by different physicalqualities (% polymer, % cross-linking of polymer, chemical compositionof scaffold, pore size, porosity, and pore architecture, stiffness,toughness, ductility, viscoelasticity, and or composition of bioactivesubstances such as growth factors, homing/migration factors,differentiation factors. Each niche has a specific effect on a cellpopulation, e.g., promoting or inhibiting a specific cellular function,attraction, adhesion, elaboration of secreted factors or enzymes, orinduction of cell death. Cells implanted within the scaffold prior toadministration of the device to a subject are engineered to targetspecific cell populations only, and permitted to migrate out of thescaffold to eliminate a target tissue. For example, engineered virusesand activated macrophages are released from the scaffold composition toeither infect target cells or clear debris from the surrounding tissue.For some purposes, the device acts locally, and is therefore placed ortransplanted into, on, or proximal to a target tissue, wherein thattarget tissue resides either internal or external (skin surfaces) to thebody. Alternatively, the device acts systemically and is thereforeintroduced into the gastrointestinal tract, blood supply, lymphaticfluid, or cerebral spinal fluid. Devices are introduced into or onto abodily tissue using a variety of known methods and tools, e.g.,gelatin-coated pill, spoon, tweezers or graspers, hypodermic needle,endoscopic manipulator, endo- or trans-vascular-catheter, stereotaxicneedle, snake device, organ-surface-crawling robot (United States PatentApplication 20050154376; Ota et al., 2006, Innovations 1:227-231),minimally invasive surgical devices, surgical implantation tools, andtransdermal patches. Devices can also be assembled in place, for exampleby sequentially injecting or inserting matrix materials. Scaffolddevices are optionally recharged with cells or with bioactive compounds.

A scaffold or scaffold device is the physical structure upon which orinto which cells associate or attach, and a scaffold composition is thematerial from which the structure is made. For example, scaffoldcompositions include biodegradable or permanent materials such as thoselisted below. The mechanical characteristics of the scaffold varyaccording to the application or tissue type for which elimination issought. In the series of experiments described in the examples, apolymer formulation consisting of poly(lactide-co-glycolide) (PLG) wasused. Other preferred polymers include but are not limited topoly(lactide)s, poly(glycolide)s, poly(lactic acid)s, poly(glycolicacid)s, polyanhydrides, polyorthoesters, polyetheresters,polycaprolactones, polyesteramides, polycarbonates, polycyanoacrylates,polyurethanes, polyacrylates, and blends or copolymers of the above. Ascaffold or scaffold device is biodegradable (e.g., collagen, alginates,polysaccharides, polyethylene glycol (PEG), poly(glycolide) (PGA),poly(L-lactide) (PLA), or poly(lactide-co-glycolide) (PLGA) or permanent(e.g., silk). In one embodiment, the scaffold or scaffold devicecomprises a biocompatible polymer matrix that may optionally bebiodegradable in whole or in part. A hydrogel is one example of asuitable polymer matrix material. Examples of materials which can formhydrogels include polylactic acid, polyglycolic acid, PLGA polymers,alginates and alginate derivatives, gelatin, collagen, agarose, naturaland synthetic polysaccharides, polyamino acids such as polypeptidesparticularly poly(lysine), polyesters such as polyhydroxybutyrate andpoly-.epsilon.-caprolactone, polyanhydrides; polyphosphazines,poly(vinyl alcohols), poly(alkylene oxides) particularly poly(ethyleneoxides), poly(allylamines)(PAM), poly(acrylates), modified styrenepolymers such as poly(4-aminomethylstyrene), pluronic polyols,polyoxamers, poly(uronic acids), poly(vinylpyrrolidone) and copolymersof the above, including graft copolymers. In another embodiment, thescaffolds may be fabricated from a variety of synthetic polymers andnaturally-occurring polymers such as, but not limited to, collagen,fibrin, hyaluronic acid, agarose, and laminin-rich gels. In the case ofbiodegradable structures, the composition is degraded by physical orchemical action, e.g., level of hydration, heat or ion exchange or bycellular action, e.g., elaboration of enzyme, peptides, or othercompounds by nearby or resident cells. The consistency varies from asoft/pliable (e.g., a gel) to glassy, rubbery, brittle, tough, elastic,stiff. The structures contain pores, which are nanoporous, microporous,or macroporous, and the pattern of the pores is optionally homogeneous,heterogeneous, aligned, repeating, or random.

Alginates are versatile polysaccharide based polymers that may beformulated for specific applications by controlling the molecularweight, rate of degradation and method of scaffold formation. Couplingreactions can be used to covalently attach bioactive epitopes, such asthe cell adhesion sequence RGD to the polymer backbone. Alginatepolymers are formed into a variety of scaffold types. Injectablehydrogels can be formed from low MW alginate solutions upon addition ofa cross-linking agents, such as calcium ions, while macroporousscaffolds are formed by lyophilization of high MW alginate discs.Differences in scaffold formulation control the kinetics of scaffolddegradation. Release rates of morphogens or other bioactive substancesfrom alginate scaffolds is controlled by scaffold formulation to presentmorphogens in a spatially and temporally controlled manner. Thiscontrolled release not only eliminates systemic side effects and theneed for multiple injections, but can be used to create amicroenvironment that activates host cells at the implant site andtransplanted cells seeded onto a scaffold.

The scaffold comprises a biocompatible polymer matrix that is optionallybiodegradable in whole or in part. A hydrogel is one example of asuitable polymer matrix material. Examples of materials which can formhydrogels include polylactic acid, polyglycolic acid, PLGA polymers,alginates and alginate derivatives, gelatin, collagen, agarose, naturaland synthetic polysaccharides, polyamino acids such as polypeptidesparticularly poly(lysine), polyesters such as polyhydroxybutyrate andpoly-epsilon, poly-caprolactone, polyanhydrides; polyphosphazines,poly(vinyl alcohols), poly(alkylene oxides) particularly poly(ethyleneoxides), poly(allylamines)(PAM), poly(acrylates), modified styrenepolymers such as poly(4-aminomethylstyrene), pluronic polyols,polyoxamers, poly(uronic acids), poly(vinylpyrrolidone) and copolymersof the above, including graft copolymers.

Scaffold Fabrication

Scaffolds are fabricated from a variety of synthetic polymers andnaturally-occurring polymers such as, but not limited to, collagen,fibrin, hyaluronic acid, agarose, and laminin-rich gels. One preferredmaterial for the hydrogel is alginate or modified alginate material.Alginate molecules are comprised of (1-4)-linked β-D-mannuronic acid (Munits) and a L-guluronic acid (G units) monomers, which can vary inproportion and sequential distribution along the polymer chain. Alginatepolysaccharides are polyelectrolyte systems which have a strong affinityfor divalent cations (e.g. Ca⁺², Mg⁺², Ba⁺²) and form stable hydrogelswhen exposed to these molecules. See Martinsen A., et al., Biotech. &Bioeng., 33 (1989) 79-89.) For example, calcium cross-linked alginatehydrogels are useful as a matrix for other cell types. cross-linkedalginate hydrogels have been used in many biomedical applications,including materials for dental impressions (Hanks C. T., et al.,Restorative Dental Materials; Craig, R. G., ed., Ninth Edition, Mosby(1993)), wound dressings (Matthew I. R. et al., Biomaterials, 16 (1995)265-274), an injectable delivery medium for chondrocyte transplantation(Atala A., et al., J Urology, 152 (1994) 641-643), and an immobilizationmatrix for living cells (Smidsrod O., et al, TIBTECH 8 (1990) 71-78).

An exemplary device utilizes an alginate or other polysaccharide of arelatively low molecular weight, preferably of size which, afterdissolution, is at the renal threshold for clearance by humans, e.g.,the alginate or polysaccharide is reduced to a molecular weight of 1000to 80,000 daltons. Preferably, the molecular mass is 1000 to 60,000daltons, particularly preferably 1000 to 50,000 daltons. It is alsouseful to use an alginate material of high guluronate content since theguluronate units, as opposed to the mannuronate units, provide sites forionic cross-linking through divalent cations to gel the polymer. U.S.Pat. No. 6,642,363, incorporated herein by reference discloses methodsfor making and using polymers containing polysaccharides such asalginates or modified alginates.

Useful polysaccharides other than alginates include agarose andmicrobial polysaccharides such as those listed in the table below.

Polysaccharide Scaffold Compositions Polymers^(a) Structure FungalPullulan (N) 1,4-; 1,6-α-D-Glucan Scleroglucan (N) 1,3; 1,6-α-D-GlucanChitin (N) 1,4-β-D-Acetyl Glucosamine Chitosan (C)1,4-β•-D-N-Glucosamine Elsinan (N) 1,4-; 1,3-α-D-Glucan BacterialXanthan gum (A) 1,4-β•-D-Glucan with D-mannose; D-glucuronic Acid asside groups Curdlan (N) 1,3-β•-D-Glucan (with branching) Dextran (N)1,6-α-D-Glucan with some 1,2; 1,3-; 1,4-α-linkages Gellan (A)1,4-β•-D-Glucan with rhamose, D-glucuronic acid Levan (N)2,6-β-D-Fructan with some β-2,1-branching Emulsan (A)Lipoheteropolysaccharide Cellulose (N) 1,4-β-D-Glucan ^(a)N—neutral, A =anionic and C = cationic.

The scaffolds of the invention are porous or non-porous. For example,the scaffolds are nanoporous having a diameter of less than about 10 nm;microporous wherein the diameter of the pores are preferably in therange of about 100 nm-20 μm; or macroporous wherein the diameter of thepores are greater than about 20 μm, more preferably greater than about100 μm and even more preferably greater than about 400 μm. In oneexample, the scaffold is macroporous with aligned pores of about 400-500μm in diameter. The preparation of polymer matrices having the desiredpore sizes and pore alignments are described in the Examples. Othermethods of preparing porous hydrogel products are known in the art.(U.S. Pat. No. 6,511,650 incorporated herein by reference).

Bioactive Compositions

The device includes one or more bioactive compositions. Bioactivecompositions are purified naturally-occurring, synthetically produced,or recombinant compounds, e.g., polypeptides, nucleic acids, smallmolecules, or other agents. The compositions described herein arepurified. Purified compounds are at least 60% by weight (dry weight) thecompound of interest. Preferably, the preparation is at least 75%, morepreferably at least 90%, and most preferably at least 99%, by weight thecompound of interest. Purity is measured by any appropriate standardmethod, for example, by column chromatography, polyacrylamide gelelectrophoresis, or HPLC analysis.

The bioactive composition affects cell mobility and survival by inducingchanges in the surface protein expression or interaction, orintracellular signaling of a cell, by restricting its access toenvironmental cues, or by contacting a cell with a modified cell typewith means to induce cell death, compromise the structural integrity ofthe target cell, or engulf the target cell. For example, at least onecell adhesion molecule is incorporated into or onto the polymer matrix.Such molecules are incorporated into the polymer matrix prior topolymerization of the matrix or after polymerization of the matrix.Examples of cell adhesion molecules include but are not limited topeptides, proteins and polysaccharides. More specifically, cell adhesionmolecules include fibronectin, laminin, collagen, thrombospondin 1,vitronectin, elastin, tenascin, aggrecan, agrin, bone sialoprotein,cartilage matrix protein, fibronogen, fibrin, fibulin, mucins, entactin,osteopontin, plasminogen, restrictin, serglycin, SPARC/osteonectin,versican, von Willebrand Factor, polysaccharide heparin sulfate,connexins, collagen, RGD (Arg-Gly-Asp) and YIGSR (Tyr-Ile-Gly-Ser-Arg)(SEQ ID NO: 9) peptides and cyclic peptides, glycosaminoglycans (GAGs),hyaluronic acid (HA), condroitin-6-sulfate, integrin ligands, selectins,cadherins and members of the immunoglobulin superfamily. Other examplesinclude neural cell adhesion molecules (NCAMs), intercellular adhesionmolecules (ICAMs), vascular cell adhesion molecule (VCAM-1),platelet-endothelial cell adhesion molecule (PECAM-1), L1, and CHL1.

Particularly preferred cell adhesion molecules of the invention arepeptides or cyclic peptides containing the amino acid sequencearginine-glycine-aspartic acid (RGD) which is known as a cell attachmentligand and found in various natural extracellular matrix molecules. Apolymer matrix with such a modification provides cell adhesionproperties to the scaffold of the invention, and sustains long-termsurvival of mammalian cell systems, as well as supporting cell growthand differentiation.

Examples of some of these molecules and their function are shown in thefollowing table.

ECM Proteins and peptides and role in cell function Seq. ProteinSequence ID No: Role Fibronectin RGDS 12 Adhesion LDV Adhesion REDV 13Adhesion Vitronectin RGDV 14 Adhesion Laminin A LRGDN 7 Adhesion IKVAV 8Neurite extension Laminin B1 YIGSR 9 Adhesion of many cells, via 67 kDlaminin receptor PDSGR 10 Adhesion Laminin B2 RNIAEIIKDA 11 Neuriteextension Collagen 1 RGDT 15 Adhesion of most cells DGEA 16 Adhesion ofplatelets, other cells Thrombospondin RGD Adhesion of most cells VTXG 17Adhesion of platelets

-   Hubbell, J A (1995): Biomaterials in tissue engineering.    Bio/Technology 13:565-576. One-letter abbreviations of amino acids    are used, X stands for any amino acid.

Additional examples of suitable cell adhesion molecules are shown below.

Amino acid sequences specific for proteoglycanbinding from extracellular matrix proteins SEQ. ID. SEQUENCE NO. PROTEINXBBXBX* 2 Consensus PRRARV 3 Fibronectin YEKPGSPPREVVPRPRPGV 4Fibronectin RPSLAKKQRFRHRNRKGYRSQRGHSRGR 5 Vitronectin RIQNLLKITNLRIKFVK6 Laminin

Coupling of the cell adhesion molecules to the polymer matrix isaccomplished using synthetic methods which are in general known to oneof ordinary skill in the art and are described in the examples.Approaches to coupling of peptides to polymers are discussed in Hiranoand Mooney, Advanced Materials, p. 17-25 (2004). Other useful bondingchemistries include those discussed in Hermanson, BioconjugateTechniques, p. 152-185 (1996), particularly by use of carbodiimidecouplers, DCC and DIC (Woodward's Reagent K). Since many of the celladhesion molecules are peptides, they contain a terminal amine group forsuch bonding. The amide bond formation is preferably catalyzed by1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC), which is a watersoluble enzyme commonly used in peptide synthesis. The density of celladhesion ligands, a critical regulator of cellular phenotype followingadhesion to a biomaterial. (Massia and Hubbell, J. Cell Biol.114:1089-1100, 1991; Mooney et al., J. Cell Phys. 151:497-505, 1992; andHansen et al., Mol. Biol. Cell 5:967-975, 1994) can be readily variedover a 5-order of magnitude density range.

Device Construction

The scaffold structure is constructed out of a number of differentrigid, semi-rigid, flexible, gel, self-assembling, liquid crystalline,or fluid compositions such as peptide polymers, polysaccharides,synthetic polymers, hydrogel materials, ceramics (e.g., calciumphosphate or hydroxyapatite), proteins, glycoproteins, proteoglycans,metals and metal alloys. The compositions are assembled into cellscaffold structures using methods known in the art, e.g., injectionmolding, lyophilization of preformed structures, printing,self-assembly, phase inversion, solvent casting, melt processing, gasfoaming, fiber forming/processing, particulate leaching or a combinationthereof. The assembled devices are then implanted or administered to thebody of a subject.

The device is assembled in vivo in several ways. The scaffold is madefrom a gelling material, which is introduced into the body in itsungelled form where it gels in situ. Exemplary methods of deliveringdevice components to a site at which assembly occurs include injectionthrough a needle or other extrusion tool, spraying, painting, or methodsof deposit at a tissue site, e.g., delivery using an application deviceinserted through a cannula. In one example, the ungelled or unformedscaffold material is mixed with bioactive substances and/or cells priorto introduction into the body or while it is introduced. The resultantin vivo/in situ assembled scaffold contains a mixture of thesesubstances and/or cells.

In situ assembly of the scaffold occurs as a result of spontaneousassociation of polymers or from synergistically or chemically catalyzedpolymerization. Synergistic or chemical catalysis is initiated by anumber of endogenous factors or conditions at or near the assembly site,e.g., body temperature, ions or pH in the body, or by exogenous factorsor conditions supplied by the operator to the assembly site, e.g.,photons, heat, electrical, sound, or other radiation directed at theungelled material after it has been introduced. The energy is directedat the scaffold material by a radiation beam or through a heat or lightconductor, such as a wire or fiber optic cable or an ultrasonictransducer. Alternatively, a shear-thinning material, such as anamphiphile, is used which re-cross links after the shear force exertedupon it, for example by its passage through a needle, has been relieved.

Suitable hydrogels for both in vivo and ex vivo assembly of scaffolddevices are well known in the art and described, e.g., in Lee et al.,2001, Chem. Rev. 7:1869-1879. The peptide amphiphile approach toself-assembly assembly is described, e.g., in Hartgerink et al., 2002,Proc. Natl. Acad. Sci. U.S.A. 99:5133-5138. A method for reversiblegellation following shear thinning is exemplified in Lee et al., 2003,Adv. Mat. 15:1828-1832

A multiple compartment device is assembled in vivo by applyingsequential layers of similarly or differentially doped gel or otherscaffold material to the target site. For example, the device is formedby sequentially injecting the next, inner layer into the center of thepreviously injected material using a needle, forming concentricspheroids. Non-concentric compartments are formed by injecting materialinto different locations in a previously injected layer. A multi-headedinjection device extrudes compartments in parallel and simultaneously.The layers are made of similar or different scaffolding compositionsdifferentially doped with bioactive substances and different cell types.Alternatively, compartments self-organize based on theirhydro-philic/phobic characteristics or on secondary interactions withineach compartment.

Compartmentalized Device

In certain situations, a device containing compartments with distinctchemical and/or physical properties is useful. Such a configuration isparticularly useful when separating quarantined cell populationsundergoing elimination from the outside tissue. For instance, theprocess or byproduct of cell destruction could interfere with theability of non-target cells to pass through the device and remainunaffected. Therefore, it is advantageous to have multiple compartments,some of which serve to contain harmful agents or cells from gainingaccess to healthy tissue. Alternatively, the device captures multipleundesirable cell types and sorts them into different chambers of thedevice where these cell populations contact bioactive compositionscontaining unique combinations of attractive, adhesive, and destructiveagents.

A compartmentalized device is designed and fabricated using differentcompositions or concentrations of compositions for each compartment. Forexample, and engineered cell population with means to induce death oftarget cells is encapsulated within hydrogels, using standardencapsulation techniques (e.g., alginate microbead formation). Thisfirst hydrogel contains factors required to maintain and contact theseengineered cells with target cells, either by their covalent coupling tothe polymer forming the gel or by their slow and sustained release fromthe gel. This compartment is then coated with a second layer of gel(e.g., double layered alginate microbeads) that contains factors that donot support the survival of these engineered cells, but instead adheretarget cells in order to expose them to the engineered population in acontrolled manner. This second compartment is formed from the samematerial that contains distinct factors (e.g., attractive and/oradhesive ligands), the same material in a distinct form (e.g., varyingmechanical properties or porosity), or a completely different materialthat provides appropriate chemical/physical properties.

Alternatively, the compartments are fabricated individually, and thenadhered to each other (e.g., a “sandwich” with an inner compartmentsurrounded on one or all sides with the second compartment). This latterconstruction approach is accomplished using the intrinsic adhesivenessof each layer for the other, diffusion and interpenetration of polymerchains in each layer, polymerization or cross-linking of the secondlayer to the first, use of an adhesive (e.g., fibrin glue), or physicalentrapment of one compartment in the other. The compartmentsself-assemble and interface appropriately, either in vitro or in vivo,depending on the presence of appropriate precursors (e.g., temperaturesensitive oligopeptides, ionic strength sensitive oligopeptides, blockpolymers, cross-linkers and polymer chains (or combinations thereof),and precursors containing cell adhesion molecules that allowcell-controlled assembly). Multiple compartments are designed to captureand either contain or eliminate target cells appropriately. In addition,the device is designed to have a number of compartments, in which cellsenter in parallel, in contrast to serially passing through allcompartments. The different compartments each capture target cellsinduce cell death of the contained cells by a unique mechanism, and inthis manner provide a means to either remove the same target cell typeby a variety of methods or to remove multiple target cell types byunique methods most appropriate for each captured population. Anindividual with ordinary skill in the art of medicine and biomaterialscan readily derive a number of potentially useful designs for a givenstarting cell type and desired daughter cell output.

Alternatively, the compartmentalized device is formed using a printingtechnology. Successive layers of a scaffold precursor doped withbioactive substances and/or cells is placed on a substrate then crosslinked, for example by self-assembling chemistries. When the crosslinking is controlled by chemical-, photo- or heat-catalyzedpolymerization, the thickness and pattern of each layer is controlled bya masque, allowing complex three dimensional patterns to be built upwhen un-cross-linked precursor material is washed away after eachcatalyzation. (W T Brinkman et al., Photo-cross-linking of type 1collagen gels in the presence of smooth muscle cells: mechanicalproperties, cell viability, and function. Biomacromolecules, 2003July-August; 4(4): 890-895; W. Ryu et al., The construction ofthree-dimensional micro-fluidic scaffolds of biodegradable polymers bysolvent vapor based bonding of micro-molded layers. Biomaterials, 2007February; 28(6): 1174-1184; Wright, Paul K. (2001). 21st Centurymanufacturing. New Jersey: Prentice-Hall Inc.) Complex,multi-compartment layers are also built up using an inkjet device which“paints” different doped-scaffold precursors on different areas of thesubstrate. Julie Phillippi (Carnegie Mellon University) presentation atthe annual meeting of the American Society for Cell Biology on Dec. 10,2006; Print me a heart and a set of arteries, Aldhouse P., New Scientist13 Apr. 2006 Issue 2547 p 19; Replacement organs, hot off the press, C.Choi, New Scientist, 25 Jan. 2003, v 2379. These layers are built-upinto complex, three dimensional compartments. The device is also builtusing any of the following methods: Jetted Photopolymer, Selective LaserSintering, Laminated Object Manufacturing, Fused Deposition Modeling,Single Jet Inkjet, Three Dimensional Printing, or Laminated ObjectManufacturing.

Incorporation of Compositions into/onto a Scaffold Device

Bioactive substances that influence attraction, adhesion, and cell deathare introduced into or onto the scaffold structures. Cell-cell adhesionmolecules (cadherins, integrins, ALCAM, NCAM, proteases) are optionallyadded to the scaffold composition.

The release profiles of bioactive substances from scaffold devices iscontrolled by both diffusion and polymer degradation, the dose of thecompound loaded in the system, and the composition of the polymer.Similarly, the range of action (tissue distribution) and duration ofaction, or spatiotemporal gradients of these substances are regulated bythese variables. The diffusion and degradation of the factors in thetissue of interest is optionally regulated by chemically modifying thefactors, or by an activity limiting factor of the substance itself,i.e., the half-life of radioisotope limits the duration of its emissioninto nearby tissue.

The bioactive substances are added to the scaffold compositions usingknown methods including surface absorption, physical immobilization,e.g., using a phase change to entrap the substance in the scaffoldmaterial. For example, a growth factor is mixed with the scaffoldcomposition while it is in an aqueous or liquid phase, and after achange in environmental conditions (e.g., pH, temperature, ionconcentration), the liquid gels or solidifies thereby entrapping thebioactive substance. Alternatively, covalent coupling, e.g., usingalkylating or acylating agents, is used to provide a stable, long-termpresentation of a bioactive substance on the scaffold in a definedconformation. Exemplary reagents for covalent coupling of suchsubstances are provided in the table below.

Methods to covalently couple peptides/proteins to polymers ReactingFunctional groups on Group of proteins/ Polymer Coupling reagents andcross-linker peptides —OH Cyanogen bromide (CNBr) —NH₂ Cyanuric chloride4-(4,6-Dimethoxy-1,3,5-triazin- 2-yl)-4-methyl- morpholinium chloride(DMT-MM) —NH₂ Diisocyanate compounds —NH₂ Diisothoncyanate compounds.—OH Glutaraldehyde Succinic anhydride —NH₂ Nitrous Acid —NH₂Hydrazine + nitrous acid —SH —Ph—OH —NH₂ Carbodiimide compounds (e.g.,EDC, —COOH DCC)[a] DMT-MM —COOH Thionyl chloride —NH₂N-hydroxysuccinimide N-hydroxysulfosuccinimide + EDC —SH Disulfidecompound —SH [a]EDC: 1-ethyl-3-(3-dimethylaminopropyl) carbodiimidehydrochloride; DCC: dicyclohexylcarbodiimide

Bioactive substances are capable of attracting target cell types towardor into the device and adhering target cell types upon contact. Otherpreferred bioactive substances are capable inducing cell death ofcaptured cells, or target cell types in close proximity of the device.Such bioactive substances are used alone or in combination to achievethe desired result.

EXAMPLES Example 1: Collection and Removal of Microorganisms to Prevent,Diagnose, or Treat Infectious Disease

Hosts exposed to or infected by invasive microorganisms such as viruses,bacteria, protozoa, fungi, or worms, typically do not present symptomsof infection immediately. By the time the host presents outward signs ofan infection, ingested microorganisms have likely matured, multiplied,spread, or inflicted damage upon host tissues. The subject is identifiedas planning to be located in an environment characterized by thepresence of such a pathogen or having recently been located in such anenvironment. The scaffold composition is administered to the hostsubject either prior to or following exposure to these microorganisms,wherein the device captures microorganism by filtration or specificadhesion proteins contained within a bioactive composition. When thedevice is administered very soon after exposure, the device eliminatesmicroorganisms, and prevents the spread of infection. Alternatively, thedevice is administered and collected in order to identify the infectiousagent to which the subject was exposed prior to the emergence ofsymptoms, providing a valuable head-start on diagnosing and treatingfuture disease. Moreover, the scaffold is administered to non-humanspecies residing in areas of suspected infestation in order to identifypotential threats to humans living in nearby as a preventative measure.

The impact of this diagnostic innovation for microorganisms can only befully appreciated in light of the number of microorganisms that cancause disease. Exemplary viral classes that infect human hosts include,but are not limited to: adenoviruses, herpesviruses, poxviruses,parvoviruses, papovaviruses, hepadnaviruses, orthomyxoviruses,paramyxoviruses, coronaviruses, picornaviruses, reoviruses, togaviruses,flaviviruses, arenaviruses, rhabdoviruses, and retroviruses. The commonnames for some of the diseases caused by the viral classes listed aboveinclude: smallpox, fifth disease, HPV, HBV, kidney disease, influenza,mumps, measles, subacute sclerosing panenephalitis, RSV, cold,poliomyelitis, hand-foot-and-mouth disease, nonspecific febrile illness,croup, bronchitis, pneumonia, hepatitis, pancreatitis, arthritis,diabetes, meningitis, encephalitis, paralysis, viral myocarditis, HAV,gastroenteritis, diarrhea, respiratory distress, rubella, respiratorytract, yellow and dengue Fever, aseptic meningitis, acute viralhaemorrhagic illness, rabies, leukemia, HTLV-I-associated myelopathy,and AIDS.

Exemplary bacterial strains that infect human hosts include, but are notlimited to: Staphylococcus aureus, Streptococcus pneumoniae,Streptococcus pyogenes, Neisseria gonorrhoeae, Neisseria meningitides,Corynebacteria diphtheriae, Bacillus anthracis, Listeria monocytogenes,Escherichia coli, Salmonella typhimurium, Salmonella typhae, Shigella,Campylobacter, Vibrio chloerae, Yersinia pestis, Pasteurella,Pseudomonas aeruginosa, Brucella Haemophilus, influenzae, Legionella,Bordetella, Clostridium tetani, Clostridium botulinum, Clostridiumperfringens, Treponema pallidum, Borrelia burgdorferi, Leptospirainterrogans, Mycobacterium tuberculosis, Mycobacterium leprae,Mycobacterium avium, Rickettsia prowazekii, Chlamydia trachomatis, andMycoplasma pneumoniae. The common names for some of the diseases orconditions caused by the bacteria listed above are: toxic shocksyndrome, pneumonia, acute sinusitis, otitis media, meningitis,osteomyelitis, septic arthritis, endocarditis, peritonitis,pericarditis, cellulitis, brain abscess, tonsilitis, scarlet fever,glomerulonephritis, rheumatic fever, gonorrhoea, meningitis, diphtheria,anthrax, listeriosis, sepsis, gastroenteritis, typhoid fever, diarrhea,anorexia, salmonellosis, bacillary dysentery, Guillain-Barré syndrome,cholera, bubonic plague, cellulitis, wound infection, brucellosis,meningitis, bacteremia, cellulitis, osteomyelitis, epiglottitis, jointinfection, sinusitis, Legionnaires' disease, pertussis or whoopingcough, tetanus, botulism, tissue necrosis, bacteremia, emphysematouscholecystitis, gas gangrene, syphilis, kidney damage, Lyme arthritis,Lyme disease, tuberculosis, leprosy, typhus, rheumatoid arthritis,prostatitis, epididymitis, cervicitis, urethritis, and infertility.

Protozoa that infect human host include, but are not limited to:Entamoeba histolytica, Giardia, Leishmania, Plasmodium (falciparum,vivax), Trypanosoma (brucei), Toxoplasma gondii, and Cryptosporidium.Diseases/conditions caused by the protozoa listed above include, but arenot limited to: Giardiasis, cutaneous (localized and diffuse)infections, visceral infections, fever, swelling of the liver andspleen, anemia, Malaria, sleeping sickness, Toxoplasmosis, andCryptosporidiosis.

Fungi that infect human hosts include, but are not limited to: Candidaalbicans, Cryptococcus neoformans, Aspergillus, Histoplasma capsulatum,Coccidioides immitis, and Pneumocystis carinii. Diseases or conditionscaused by the fungi listed above are: Candidiasis; skin, lung, andmeningeal infections; fever, cough, chest pain or breathlessness;Histoplasmosis; Coccidioidomycosis (Valley Fever), and Pneumonia.

Worms that infect human hosts include, but are not limited to: Trichuristrichiura, Trichinella spiralis, Enterobius vermicularis, Ascarislumbricoides, Ancylostoma, Strongyloides, Filaria, Onchocerca volvulus,Loa loa, Dracuncula medinensis, Schistosoma (mansoni), and Clonorchissinensis. Diseases or conditions caused by worms listed above include,but are not limited to: human whipworm, roundworm, trichuriasis,Trichinosis, Pinworm, Ascariasis, Hookworm; intellectual, cognitive andgrowth retardation; intrauterine growth retardation, prematurity, andlow birth weight; elephantiasis, Lymphatic Filariasis; river blindness;Calabar swellings; Dracunculiasis/Guinea worm disease (GWD);Schistosomiasis; liver and intestinal damage.

For example, the scaffold composition is packaged into a gelatin-coatedpill and administered orally, releasing a multiple number of scaffoldcompositions within the gastrointestinal tract of the host subject. Fordiagnosis, collection of the scaffolds occur using a stool sample fromthe subject. This method of internalizing and collecting the deviceavoids exposure of the host and administrator to blood, which isparticularly useful in regions of the world where blood-born viralinfections, like AIDS, are prevalent. The device is alternativelyintroduced by intravenous, intraperitoneal, or intramuscular injections,as well as lumbar puncture, subcutaneous implantation, or surgicalmethods depending on the target site of interest.

Example 2: Capture and Collection of Circulating Mammalian Cells toDiagnose Disease

The device is also used to diagnose disease caused by mammalian oreukaryotic cells, whereby it is administered into a host and collected.Following collection, the device is manipulated to yield its contents inorder to allow for the identification of undesirable mammalian cellsthat circulate within the host, such as cancer cells undergoingmetastasis from their primary tumor location, leukemia, lymphoma,sickle-shaped blood cells, or immune cells contributing to inflammation.Often a subject can be afflicted with primary cancer tumors affectingmultiple organs all of which could metastasize and travel through theblood stream, lymphatic fluid, or cerebral spinal fluid (CSF). Toquickly diagnose which tumor is contributing to the metastasis, and toavoid the unnecessary risk of surgery or multiple biopsies, the deviceis used to collect these cells for identification based either onmolecular expression profiles or cell morphological features.

Example 3: Elimination of Neoplasms by Implantation of a BiodegradableDevice

To counteract the growth of either malignant or benign neoplasms, whichcould become malignant, the device is implanted either by subcutaneousor intraperitoneal injection, or by surgical implantation, at the siteof abnormally rapid cell replication. Once in place, the scaffoldcomposition adheres to adjacent tissue and a bioactive composition thatis either tethered to the scaffold or allowed to disseminate from thescaffold at close range, induces death of the rapidly dividing cellpopulation. In this case, the scaffold degrades over time and does notrequire collection. A significant advantage of using the scaffoldcomposition to reduce neoplastic growth, is that devices located aroundthe tumor borders capture cells that attempt to escape from the tumor,thus, preventing them from spreading outside of their local niche.

Example 4: To Remove Blockages from Arterial Walls Using the Device as aStent

To combat high blood pressure and minimize the risk of stroke, thedevice is introduced into the circulatory system by one of two methods:the scaffold is either used as an arterial stent in order to eitherremain stationary and allow the blood to pass through, or as a mobiledevice that flows along with blood in order to gain access to the entirebody. As an arterial stent, the device reduces or eliminates theaccumulation of compounds and/or cells that threaten to block the normalflow of blood. As a mobile device, the scaffold composition prevents thefuture occlusion of arteries by eliminating these compounds and/or cellsbefore they can attach to the arterial walls.

Example 5: Device Implantation for the Permanent Removal of Adipocytesto Accomplish Life-Long Weight Loss

Another use for the scaffold composition is “surgery-less liposuction.”Many subjects are incapable of overcoming morbid obesity due to what isnow considered a chemical addition to food. Methods to alter lifestyleand surgical intervention to limit the capacity of the body to intakecalories have met with limited success. However, the elimination ofadipocytes, or fat cells, from the body by means of destroying themlocally, is more permanent solution the success of which does not dependupon the subject to maintain a proper diet. This therapeutic strategyrelieves the chemical and physical threats that inflated adipocytes poseon the internal organs of morbidly obese subjects. To reduce oreliminate adipocytes, the scaffold composition is delivered bysubcutaneous or intraperitoneal injection, or surgical implantation,into target sites where it captures or adheres to, and subsequentlydestroys adipocytes until the scaffold itself degrades.

Example 6: Prevention of Glial Scars that Physically Prohibit CentralNerve Regeneration

A unique property of the central nervous system, as opposed to theperipheral nervous system, is the inhibition of nerve regeneration.While researchers have identified the signaling molecules that preventcentral nerve re-growth, Nogo ligands and their receptors, anotherfactor that threatens the success of regenerative treatments is glialscarring. Glial cells, which normally support neuronal function, respondto nerve injury by increasing replication and invading the site ofdamage, thus forming a physical blockade against nerve outgrowth. Thescaffold composition is delivered to the site of central nerve injury,and a bioactive composition is incorporated having means to specificallyadhere, capture, and destroy glial cells that accumulate close to thesite of nerve injury. Thus, the device prevents a glial scar fromimpeding regeneration. Furthermore, inhibitors of Nogo or blockingagents of the Nogo receptor diffuse from a bioactive composition furtherincorporated into the scaffold composition to eliminate molecularsignals that would also inhibit regeneration.

Example 7: In Vivo Clearance of Amyloid Protein Plaques Associated withAlzheimer's Disease

Current research into the field of neurodegenerative disease hasprovided important insights into the mechanisms behind some of our mostdebilitating diseases. For example, Alzheimer's Disease (AD) istentatively diagnosed in living subjects based primarily on behavioralaffects and, in some cases, genetic tests which can show predispositiononly. The final diagnosis for AD is made postmortem, and the presence ofprotein plaques on the brain provides the final confirmation. Todiagnose, retard, or arrest the formation of amyloid plaques in thebrains of suspected AD patients, the device is injected into thecerebral spinal fluid by lumbar puncture wherein the scaffoldcomposition contains immune cells capable of clearing cellular debris,and wherein the scaffold composition adheres to amyloid protein, thusallowing these resident cells to clear AD-related depositions. Thedevise is constructed of a non-biodegradable material and remains in thecentral nervous system to eliminate future deposits from accumulating.

Example 8: Prevention or Inhibition of Autoimmune Disease

Autoimmune disorders, e.g. those of the central nervous system arecharacterized by neurodegeneration and inappropriate immune systemresponses. For instance, multiple sclerosis is an autoimmune conditionin which the host's immune system incorrectly recognizes nerve fibers asforeign cells, leading to their demyelination and, ultimately, to deathof the host. To retard or arrest the progression of MS, the device isinjected into the cerebral spinal fluid via lumbar puncture, orcontacted to either brain or spinal cord tissue by surgical methods,wherein a bioactive composition incorporated into the scaffoldcomposition, containing self-antigens (pre-isolated from the givensubject) and chemokines, attracts, captures, and eliminates immune cellsthat contribute to immune system attacks of host tissues.

Other Embodiments

While the invention has been described in conjunction with the detaileddescription thereof, the foregoing description is intended to illustrateand not limit the scope of the invention, which is defined by the scopeof the appended claims. Other aspects, advantages, and modifications arewithin the scope of the following claims.

The patent and scientific literature referred to herein establishes theknowledge that is available to those with skill in the art. All UnitedStates patents and published or unpublished United States patentapplications cited herein are incorporated by reference. All publishedforeign patents and patent applications cited herein are herebyincorporated by reference. All other published references, documents,manuscripts and scientific literature cited herein are herebyincorporated by reference.

While this invention has been particularly shown and described withreferences to preferred embodiments thereof, it will be understood bythose skilled in the art that various changes in form and details may bemade therein without departing from the scope of the inventionencompassed by the appended claims.

What is claimed is:
 1. A method of removing or eliminating a cancer cellfrom a mammalian subject, comprising a) administering to the subject, adevice comprising a scaffold composition and a bioactive composition,said bioactive composition being incorporated into or conjugated ontosaid scaffold composition, wherein said scaffold composition comprises apolymer matrix and pores, and wherein the bioactive compositioncomprises a cell attractant and a cell death-inducing agent, whereinsaid scaffold composition does not comprise cells prior to saidadministering step, b) exposing said cancer cell to said cellattractant, thereby recruiting the cancer cell to the scaffoldcomposition, c) capturing said cancer cell within said scaffoldcomposition, thereby sequestering said cancer cell from the hostenvironment, and d) killing the sequestered cancer cell within saidscaffold composition by exposure to the scaffold composition or to thecell death-inducing agent, wherein the sequestered cancer cell is killedwithout removing the device from the subject, thereby removing oreliminating the cancer cell from the subject.
 2. The method of claim 1,wherein the cell attractant disseminates from said scaffold composition.3. The method of claim 1, wherein said device is administered orally,systemically, by lumbar puncture to access cerebral spinal fluid,subcutaneously, transcutaneously, as an arterial stent, or surgically.4. The method of claim 1, further comprising the step of collecting saiddevice from the subject.
 5. The method of claim 1, wherein the bioactivecomposition comprises an antigen or an adhesive compound.
 6. The methodof claim 1, wherein the cancer cell comprises a metastatic cancer cell.7. The method of claim 1, wherein the cancer cell displays a surfaceantigen.
 8. The method of claim 1, wherein the cell attractant comprisesa cytokine or a chemokine.
 9. The method of claim 8, wherein thecytokine or chemokine attracts metastatic cancer cells, immune cells, orboth metastatic cancer cells and immune cells.
 10. The method of claim1, wherein the bioactive composition comprises an inhibitor of chemokinereceptor activity.
 11. The method of claim 10, wherein the inhibitorcomprises AMD3100, T22, TN14003, T140, TC14012, 4F-bTE, BX-471,Met-CCL5, TAK779, or TAK220.
 12. The method of claim 1, wherein thebioactive composition comprises an anti-metastatic compound.
 13. Themethod of claim 1, wherein the device adheres to a surface protein onthe cancer cell, wherein the surface protein is unique to cancer cells.14. The method of claim 1, wherein the polymer comprisespoly(lactide-co-glycolide)(PLGA).
 15. The method of claim 1, wherein thesize of the pores is less than about 10 nm, in the range of 100 nm to 2μM, or greater than about 20 μM in diameter.
 16. The method of claim 15,wherein the size of the pores is about 400 μM to 500 μM in diameter. 17.The method of claim 1, wherein the cell death-inducing agent comprises acompound that prevents or arrests cell division, an engineered virus, ahumanized monoclonal antibody to activate the immune system, apro-apoptotic ligand, a radioactive isotope, a toxin, or achemotherapeutic agent.
 18. The method of claim 17, wherein thepro-apoptotic ligand comprises FasL, TNF, TRAIL, or a caspase activator.19. The method of claim 17, wherein the radioactive isotope comprisesBismuth-213, Holmium-66, Iodine-125, Iodine-131, or Lutetium-177. 20.The method of claim 17, wherein the toxin comprises a hemotoxin, anecrotoxin, or a neurotoxin.
 21. The method of claim 17, wherein thecompound that prevents or arrests cell division comprises Dacarbazine,Fluorouracil, Fludarabine, Gemcitabine, Trastuzumab, Hydroxyurea,Idarubicin, Ifosfamide, Irinotecan, Cladribine/Leustatin,Mercaptopurine, Methotrexate, Mithramycin, Mitomycin, Mitoxanthrone,Navelbine, Nitrogen Mustard, Rituxan, Paclitaxel, Docetaxel, Topotecan,Velban, Vincristine, or Etoposide.
 22. The method of claim 1, whereinthe cell attractant comprises a multivalent acrylamide polymer linked toa sugar, a polylysine linked to a sugar, or an amino acid.
 23. Themethod of claim 22, wherein the sugar comprises galactose, fructose, orribose.
 24. The method of claim 22, wherein the amino acid comprisesaspartate, leucine, or serine.
 25. The method of claim 1, wherein thedevice is administered by implanting into a tissue of the subject whichis not blood tissue.
 26. The method of claim 25, wherein the device isadministered by implanting into a site selected from the groupconsisting of adipose tissue, gastrointestinal tract, lymphatic fluid,primary tumor site, and cerebral spinal fluid.
 27. The method of claim1, wherein the device is not collected from the subject.